Discrimination of algal and bacterial alkaline phosphatases with a differential-inhibition technique

Abstract

Both phytoplankton and bacterioplankton produce alkaline phosphatases, but the techniques currently available for discriminating between the two sources are poor, especially when samples are from turbid waters. A novel approach, based on the differential inhibition of alkaline phosphatases by various physical and chemical treatments, was assessed as a rapid and inexpensive technique for determining whether phytoplankton or bacterioplankton were the more important producers of alkaline phosphatases in turbid rivers of south-eastern Australia. Eight phytoplankton strains and 14 bacterial strains (eight isolated from the Ovens River and six isolated as bacterial contaminants of the phytoplankton cultures) were grown in laboratory culture. Rates of alkaline phosphatase activity in the bacterial cultures varied from <1 to 21 fmol cell^-1 day ^-1. Rates for phytoplankton were usually < 5µmol (µg chla) ^-1 day^-1 but could reach 128 µmol (µ chla)^-1 day^-1, depending on whether inorganic or organic phosphorus was supplied. Differential-inhibition profiles were determined for the 22 isolates, using seven chemical treatments (L-cysteine, EDTA and L-levamisole, and Zn²⁺ and Cu²⁺ each at two concentrations) plus one physical treatment (thermal deactivation). The alkaline phosphatases of the three microbial groups (i.e. Ovens River bacteria, bacteria isolated as contaminants from algal cultures, and phytoplankton) could be classified with a predictive accuracy of better than 90% when these data were analysed with Discriminant Function Analysis. L-Cysteine, Zn²⁺ and Cu²⁺ were the best predictors of class membership; thermal deactivation and EDTA sometimes were also significant. Inhibition profiles were then determined for the alkaline phosphatases of various river-water samples. These environmental samples usually (>70% of cases) grouped separately from those of the laboratory cultures of bacteria and phytoplankton, perhaps because the microbes studied in laboratory culture were not representative of native assemblages or because the culture conditions did not mimic those in nature. Nevertheless, differential-inhibition techniques have much potential for determining the origin of the alkaline phosphatases found in natural waters, with the major factor limiting their application being the collection of valid inhibition profiles for native bacterial and algal communities.